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Environmental Applications of ESI FT-ICR Mass Spectrometry

Dissertation Defense January 7, 2010. Environmental Applications of ESI FT-ICR Mass Spectrometry. Oxidized Peptide and Metal Sulfide Clusters. Jeffrey M. Spraggins University of Delaware Department of Chemistry & Biochemistry Advisor: Douglas Ridge, Ph.D. Overview.

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Environmental Applications of ESI FT-ICR Mass Spectrometry

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  1. Dissertation Defense January 7, 2010 Environmental Applications of ESI FT-ICR Mass Spectrometry Oxidized Peptide and Metal Sulfide Clusters Jeffrey M. Spraggins University of Delaware Department of Chemistry & Biochemistry Advisor: Douglas Ridge, Ph.D.

  2. Overview ESI FT-ICR Mass Spectrometry Oxidized Peptides Metal Sulfide Clusters • Johnston Group • Dr. Laskin: PNNL Luther Group (Marine Studies) • Energy-resolved SID • Molecular Dynamics • RRKM Theory Molecular Capping Agents Gas Phase Ion-Molecule Reactions Fragmentation Mechanisms Metal Sulfide Formation

  3. ESI FT-ICR Mass Spectrometry Oxidized Peptides Jeffrey M. Spraggins, Julie A. Lloyd, Murray V. Johnston, Julia Laskin, Douglas P. Ridge J.Am. Soc. Mass Spec. (2009), Accepted for puplication. Julie A. Lloyd, Jeffrey M. Spraggins, Murray V. Johnston, Julia Laskin, J. Am. Soc. Mass Spec. (2006), 17(9), 1289-1298

  4. Oxidation Reaction AngII + O Tyr* O3 AngII AngII + 3O His* (~143µM) AngII + 4O Tyr* and His*

  5. Asp-Arg-Val-Tyr-Ile-His-Pro-Phe

  6. 6T Magnet Quad Bender Lens 3 Lens 4 Lens 5 Deceleration SAM Lens 2 Lenses Surface Segmented Trapping Lenses Plates Lens 1 Accumulation Quadrupole 2 mTorr Resolving Quadrupole 510-5 Torr Collisional Quadrupole 20-50 mTorr Ion Source Ion Funnel 0.6-0.8 Torr Conductance Limit Collimating Plate Trapping Plates 6 T FT-ICR MS: PNNL Laskin J.; Denisov E. V.; Shukla A. K.; Barlow S. E.; Futrell J. H. Analytical chemistry 2002, 74, 3255-61.

  7. AngII+3O AngII Intensity AngII+4O AngII+O m/z (b-d: 43eV SID Spectra) m/z

  8. Asp-Arg-Val-Tyr-Ile-His-Pro-Phe y7

  9. Energy Resolved FECs: AngII Relative Intensity Collision Energy (eV)

  10. Acidic Amino Acids J. Laskin J. Futrell V. Wysocki J. Beauchamp Scheme Ib Scheme Ia y7

  11. AngII: charge delocalization

  12. Asp-Arg-Val-Tyr(+O)-Ile-His(+3O)-Pro-Phe

  13. Asp-Arg-Val-Tyr(+O)-Ile-His(+3O)-Pro-Phe b4+O

  14. Energy Resolved FECs: AngII+O Relative Intensity Collision Energy (eV)

  15. AngII+O b4 fragmentation pathway Scheme II b4+O

  16. AngII+O: b4 fragment

  17. Acidic Amino Acids J. Laskin J. Futrell V. Wysocki J. Beauchamp Scheme Ib Scheme Ia

  18. Asp-Arg-Val-Tyr(+O)-Ile-His(+3O)-Pro-Phe

  19. Asp-Arg-Val-Tyr(+O)-Ile-His(+3O)-Pro-Phe -45 -71 -88 b5

  20. AngII+3O [MH+3O]+-45 & [MH+3O]+-71 Scheme III [MH+3O]+-45 [MH+3O]+-71

  21. AngII+3O [MH+3O]+-88 & b5 fragmentation pathway Scheme IV [MH+3O]+-88 b5

  22. Transition States RRKM Theory Relative Intensity Collision Energy (eV)

  23. ΔS‡(cal/mol K) Entropic Considerations -25.9 -21.6 -17.0 y7 fragment b4+O fragment [AngII+3O]-45 Side Chain Backbone Amide Bond

  24. Conclusions • Unmodified Ang II • Charge-remote selectivity towards y7 fragment • M+O adduct FECs suggest that the b4 pathway is a charge-remote process. • C-term Tyr* - Backbone interaction leads to b4 fragment ion. • M+3O product FECs show both charge-remote and charge-directed selective fragmentation channels are opened with the oxidation of the His residue. • Loss of 45m/z and 71m/z driven by strong H-bonding within the His*side chain. • His* is thought to compete with Arg for the lone proton leading to the b5 and [MH+3O]+-88 charge-directed fragments. • RRKM results imply destabilization due to entropic effects. • Consistent with proposed fragmentation mechanisms for oxidation products

  25. The Big Picture • Structurally characterize peptide degradation by ozone exposure • Contribution to Fundamental MS/MS Literature • Understanding oxidized peptide fragmentation could benefit MS protein structural research • Method for increasing fragmentation sequence coverage for proteomic studies

  26. Metal Sulfide Clusters ESI FT-ICR Mass Spectrometry

  27. - Project Goal Dissolved Ions - + + • Metal Sulfide Precipitation - Cluster Aggregation† • Experimental Approach - Capping Agents - Ion-Molecule Reactions Clusters (Dissolved Polynuclear Molecular Species) 1 Nanoparticles (1-100nm) 10 ∞ † T. F. Rozan, M. E. Lassman, D. P. Ridge, G. W. Luther. Evidence for iron, copper and zinc complexation as multinuclear sulphide clusters in oxic rivers. Nature, 2000, 406, 879-882. Solid state material Size (nm)

  28. - Project Goal Dissolved Ions - + + Mass Spectrometry Clusters 1 UV/VIS Spectroscopy Nanoparticles 10 ∞ Solid state material Size (nm)

  29. 7T FT-ICR MS: UD

  30. ESI of Metal Salt Solutions Observed Nucleation Processes

  31. Salt Clusters Observed using ESI FT-ICR MS

  32. Signal Variance • [Cd(CH3COO)3]- • [Cd2(CH3COO)5]- • [Cd3(CH3COO)7]- • [Cd4(CH3COO)9]- • [Cd5(CH3COO)11]- •[Cd6(CH3COO)13]- • [Cd7(CH3COO)15]- • [Cd8(CH3COO)17]- Sampling was done using separate aliquots for each spectrum and A single aliquot was continually injected as spectra were repeatedly taken. 0.3 mM cadmium acetate water/MeOH solution (1:1)

  33. Important Points • Variance in signal over time suggests that clusters observed using ESI are forming in solution. • Results highlight processes taking place in low dielectric constant solvents • Water/MeOH (ε=66†) • Hydrothermal Fluids (ε=10-23‡) †Akerlof, G. Dielectric Constant of some Organic Solvent-Water Mixtures at various Temperatures. J. Am. Chem. Soc. 1932, 54, 4125-4139. ‡Weingartner, H.; Franck, E. U. Supercritical Water as a Solvent. Angew Chem Int Ed Engl. 2005, 44, 2672-2692.

  34. Making Metal Sulfide Clusters Molecular Capping Agents

  35. Solution Chemistry: Mononuclear Anions • 3 mM aqueous solution of Cd(NO3)2•4H2O diluted 50/50 with MeOH. • 1.5 mM aqueous solution of Cd(NO3)2•4H2O and 2-mercaptopyridine (1:1) diluted 50/50 with MeOH. • H2S(g) bubbled through a 3 mM aqueous solution of Cd(NO3)2•4H2O followed by addition of 2-mercaptopyridine (1:1), then diluted 50/50 with MeOH.

  36. Solution Chemistry: Binuclear Anions • 3 mM aqueous solution of Cd(NO3)2•4H2O diluted 50/50 with MeOH. • 1.5 mM aqueous solution of Cd(NO3)2•4H2O and 2-mercaptopyridine (1:1) diluted 50/50 with MeOH. • H2S(g) bubbled through a 3 mM aqueous solution of Cd(NO3)2•4H2O followed by addition of 2-mercaptopyridine (1:1), then diluted 50/50 with MeOH.

  37. Solution Chemistry: Trinuclear Anions • 3 mM aqueous solution of Cd(NO3)2•4H2O diluted 50/50 with MeOH. • 1.5 mM aqueous solution of Cd(NO3)2•4H2O and 2-mercaptopyridine (1:1) diluted 50/50 with MeOH. • H2S(g) bubbled through a 3 mM aqueous solution of Cd(NO3)2•4H2O followed by addition of 2-mercaptopyridine (1:1), then diluted 50/50 with MeOH.

  38. Important Points • Sulfidic clusters are observed when 2-mercaptopyridine is introduced. • Reactivity with H2S is selective based on cluster size for [Cdx(NO3)2x+1]- species. • Sampling a solution based process • Gas phase ion-molecule reactions

  39. Making Metal Sulfide Clusters Gas Phase Ion-Molecule Reactions

  40. Cadmium Salt Clusters Reactive Clusters [Cd(CH3COO)3]- [Cd2(CH3COO)5]- [Cd3(CH3COO)7]- [Cd4(CH3COO)9]- [Cd2(CH3COO)3]+ [Cd3(CH3COO)5]+ [Cd4(CH3COO)7]+ Non-Reactive Clusters [Cd(NO3)3]- [CdxCl2x+1]- [ZnxCl2x+1]- [Cd2(NO3)5]- [Cd3(NO3)7]- [Cd4(NO3)9]- [CdNO3(CH3OH)]+ [CdNO3(CH3OH)(H2O)]+ [CdCl(CH3OH)]+ [Cd2Cl3(CH3OH)]+ [ZnCl(CH3OH)]+

  41. [Cd2(NO3)5]- {536 m/z} H2S H2S -2HNO3 -2HNO3 [Cd2S(NO3)3]- [Cd2SH(NO3)4]- -HNO3 {444 m/z} {507 m/z} H2S -2HNO3 [Cd2(SH)2(NO3)3]- {478 m/z}

  42. Cd2S0 [Cd2(NO3)5]- {536 m/z} H2S H2S -2HNO3 -2HNO3 Cd2S1 [Cd2S(NO3)3]- [Cd2SH(NO3)4]- -HNO3 {444 m/z} {507 m/z} H2S -2HNO3 Cd2S2 [Cd2(SH)2(NO3)3]- {478 m/z}

  43. d[S0] S0 [Cd2(NO3)5]- = -k1[S0] r = k[Cluster Family][H2S] dt {536 m/z} k1 r = k[Cluster Family] -CH3COOH H2S • H2S is in excess • Pseudo First Order kinetics [Cd2(NO3)4SH]- d[S1] {507 m/z} = k1[S0] – k2[S2] S1 dt [Cd2S(NO3)3]- {444 m/z} k2 -CH3COOH H2S d[S2] Integrated Rate Equations [Cd2(NO3)3(SH)2]- = k2[S1] S2 {478 m/z} dt [S0]= [A]0 e-k1t k3 = 0 k1 [S1]= [A]0 (e-k1t - e-k2t) k2-k1 e-k1t e-k2t e-k3t [S2]= k1k2 + + (k2-k1)(k3-k1) (k1-k2)(k3-k2) (k1-k3)(k2-k3)

  44. [Cd2(NO3)5]- +H2S (g) 4x10-9 torr [Cd2SH(NO3)4]- [Cd2S(NO3)3]- [Cd2(SH)2(NO3)3]-

  45. [Cd2(NO3)5]- +H2S [Cd2S0]- k1= 0.084 ± 0.004s-1 [Cd2SH(NO3)4]- [Cd2S(NO3)3]- [Cd2(SH)2(NO3)3]- [Cd2S1]- k2= 0.019 ± 0.001s-1 [Cd2S2]- H2S: 4x10-9 torr

  46. Reaction Efficiency [Cd2S0]- k1= 0.084 ± 0.004s-1 • Assume fastest RXN is collision rate limited [Cd(NO3)(CH3OH)]+ k1= 2.85 ± 0.27 s-1 • Capture Collision Theory† [Cd2S1]- k2= 0.019 ± 0.001s-1 [Cd2S2]- H2S: 4x10-9 torr

  47. Reaction Efficiency [Cd2S0]- k1= 0.084 ± 0.004s-1 • Assume fastest RXN is collision rate limited [Cd(NO3)(CH3OH)]+ k1= 2.85 ± 0.27 s-1 • Capture Collision Theory† [Cd2S1]- k2= 0.019 ± 0.001s-1 [Cd2S2]- H2S: 4x10-9 torr

  48. Reaction Efficiency [Cd2S0]- k1= 0.084 ± 0.004s-1 k1/kc= 0.031 • Assume fastest RXN is collision rate limited [Cd(NO3)(CH3OH)]+ k1= 2.85 ± 0.27 s-1 • Capture Collision Theory† [Cd2S1]- k2= 0.019 ± 0.001s-1 k2/kc= 0.0069 [Cd2S2]- H2S: 4x10-9 torr

  49. Anionic Cadmium Clusters

  50. Anionic Cadmium Clusters Most Anionic Metal Cluster reactions are < 5% efficient.

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